Glucosylglycerol and proline reverse the effects of glucose on Rhodosporidium toruloides lifespan.

Rhodosporidium toruloides is a novel cell factory used to synthesis carotenoids, biosurfactants, and biofuel feedstocks. However, research on R. toruloides has generally centred on the manufacture of biochemicals, while analyses of its longevity have received scant attention. Understanding of R. toruloides longevity under different nutrient conditions could help to improve its biotechnological significance and metabolite production. Glucosylglycerol (GG) and proline are osmoprotectants that could revert the harmful effects of environmental stress. This study examined how GG and proline affect R. toruloides strain longevity under glucose nutrimental stress. Herein, we provide evidence that GG and proline enhance cell performance and viability. These compatible solutes neutralises the pro-ageing effects of high glucose (10% glucose) on the yeast cell and reverse its cellular stress. GG exhibits the greatest impact on lifespan extension at 100 mM, whereas proline exerts effect at 2 mM. Our data reveal that these compounds significantly affect the culture medium osmolarity. Moreso, GG and proline decreased ROS production and mitohormetic lifespan regulation, respectively. The data indicates that these solutes (proline and GG) support the longevity of R. toruloides at a pro-ageing high glucose culture condition.

Biorefinery of vineyard winter prunings for production of microbial lipids by the oleaginous yeast Cryptococcus curvatus.

Spent biomass from agricultural and forestry industries are substantial low-cost carbon source for reducing the input of microbial lipid production. Herein, the components of the vineyard winter prunings (VWPs) from 40 grape cultivars were analyzed. The VWPs contained (w/w) cellulose ranged from 24.8% to 32.4%, hemicellulose 9.6% to 13.8%, lignin 23.7% to 32.4%. The VWPs from Cabernet Sauvignon was processed with the alkali-methanol pretreatment, and 95.8% of the sugars was released from the regenerated VWPs after enzymatic hydrolysis. The hydrolysates from the regenerated VWPs was suitable for lipid production without further treatment as a lipid content of 59% could be achieved with Cryptococcus curvatus. The regenerated VWPs was also used for lipid production via simultaneous saccharification and fermentation (SSF), which led to a lipid yield of 0.088 g/g raw VWPs, 0.126 g/g regenerated VWPs and 0.185 g/g from the reducing sugars. This work demonstrated that the VWPs can be explored for co-production of microbial lipids.

Exogenous l-proline improved Rhodosporidium toruloides lipid production on crude glycerol.

BACKGROUND: Crude glycerol as a promising feedstock for microbial lipid production contains several impurities that make it toxic stress inducer at high amount. Under stress conditions, microorganisms can accumulate l-proline as a safeguard. Herein, l-proline was assessed as an anti-stress agent in crude glycerol media. RESULTS: Crude glycerol was converted to microbial lipids by the oleaginous yeast Rhodosporidium toruloides CGMCC 2.1389 in a two-staged culture mode. The media was supplied with exogenous l-proline to improve lipid production efficiency in high crude glycerol stress. An optimal amount of 0.5 g/L l-proline increased lipid titer and lipid yield by 34% and 28%, respectively. The lipid titer of 12.2 g/L and lipid content of 64.5% with a highest lipid yield of 0.26 g/g were achieved with l-proline addition, which were far higher than those of the control, i.e., lipid titer of 9.1 g/L, lipid content of 58% and lipid yield of 0.21 g/g. Similarly, l-proline also improved cell growth and glycerol consumption. Moreover, fatty acid compositional profiles of the lipid products was found suitable as a potential feedstock for biodiesel production. CONCLUSION: Our study suggested that exogenous l-proline improved cell growth and lipid production on crude glycerol by R. toruloides. The fact that higher lipid yield as well as glycerol consumption indicated that l-proline might act as a potential anti-stress agent for the oleaginous yeast strain.

Automated Pressurized Liquid Extraction of Microbial Lipids from Oleaginous Yeasts.

The lipids produced by oleaginous microbes are considered sustainable resources for biofuels. To facilitate controlled lipid production and lipid analysis, more efficient lipid extraction methods are required. This study describes the automated pressurized liquid extraction (APLE) method for lipid extraction from dried cells of the oleaginous yeast species Rhodosporidium toruloides and Cryptococcus curvatus. Cells were mixed with diatomite in a mortar, added to the sample chamber, and treated with a mixture of chloroform and methanol at 105 °C. More than 95% lipids were extracted. Analysis by using high-performance thin-layer chromatography showed that the neutral lipid contents in the obtained samples by APLE method were similar to those by the ball milling-assisted extraction method. The lipids had an essentially identical fatty acid composition compared with lipids extracted with the acid-heating extraction (AHE) method. This demonstrated that lipids can be efficiently extracted from oleaginous yeasts in less time and without harsh pretreatment procedures.

Utilization of Amino Acid-Rich Wastes for Microbial Lipid Production.

To produce microbial lipids for biofuel production, carbohydrates and related compounds from biomass have been routinely utilized, yet amino acids (AA) from protein-rich wastes have been overlooked so far. We use the oleaginous yeast Cryptococcus curvatus ATCC 20509 as a lipid producer and evaluate the capacity for lipid production on proteinogenic AA individually or in designated blends under two-staged culture conditions. It was found that cellular lipid contents reached 48.8%, 44.5% and 29.0% when yeast cells were cultivated in media-contained AA blends with compositional profiles similar to those of sheep viscera, meat industry by-products and fish muscle, respectively, and that lipid coefficients were more than 0.10 g g-1. Furthermore, cellular lipid contents were higher than 20% when most AA were used individually. High lipid coefficients of over 0.23 g g-1 were observed when Pro, Trp or Leu were used as a substrate. Results also indicated that higher initial media pH or reduced phosphate concentration was beneficial for lipid production on AA. This work demonstrated the potential to use AA and related wastes as substrates for microbial lipid production by the yeast C. curvatus, which fit well with the protein-based biorefinery concept. Further efforts should be devoted to recognizing the metabolic features, identifying more robust lipid producer and optimizing lipid production processes.